Huntington's disease (HD) is a progressive and fatal neurologic disorder caused by an expanded CAG repeat in the gene coding for a protein of unknown function that has been named huntingtin. Recent studies of huntingtin mRNA or protein have given the impression that it is ubiquitously expressed in neurons and so cannot itself explain selective vulnerability. When closely examined, these studies, as well and our preliminary studies, suggest that while most neurons may express huntingtin, there are significant differences in the levels of expression among particular types of neurons and between different intraneuronal compartments. We hypothesize that the neurons most susceptible to neurodegeneration in HD have especially high levels of huntingtin expression. The first goal of this proposal is to characterize the heterogeneity of huntingtin expression in rat brain at the cellular and subcellular level using quantitative immunocytochemical methods. The result will be a detailed picture of its neuronal localization and an understanding of which neuronal processes and organelles are significantly associated with huntingtin. These studies in rat will serve as a basis for our second goal in Aim l which is to characterize the differential localization of huntingtin expression in human brain tissue in distinct neuronal types and to compare controls and HD cases. In particular, we will test our hypothesis that the selectivity of neuronal death in HD can be correlated to differences in huntingtin expression. The exact cause of neuronal death in HD is unknown, however, the leading hypothesis is that of excitotoxicity induced by a defect in energy metabolism that may be caused by oxidative stress. Our second aim will examine whether cellular markers for oxidative damage (DNA fragmentation, 8-hydroxy deoxyguanosine, malondialdehyde, heme oxygenase, and 3- nitrotyrosine) are increaseD in HD brain; whether they are selectively increased in neurons with the highest levels of huntingtin expression; and whether there is a relationship between the degree of polyglutamine expansion and the degree of oxidative damage. To help determine whether oxidative damage is a primary contributor to neuronal death or a secondary result of neurodegeneration, we will study these same markers in fibroblast cell lines from HD patients and in transfected PC-12 cell lines containing a wide range of CAG repeats. These studies may help establish links between the expression and localization of huntingtin, oxidative stress, and neuronal cell death in HD.